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Independent SH2-binding sites mediate interaction of Dok-related protein with RasGTPase-activating protein and Nck

P Lock1, F Casagranda, A R Dunn

  • 1Ludwig Institute for Cancer Research and the Cooperative Research Center for Cellular Growth Factors, P. O. Box 2008, Royal Melbourne Hospital, Parkville 3050, Australia. Peter.Lock@Ludwig.edu.au

The Journal of Biological Chemistry
|July 31, 1999
PubMed

Insights

Researchers identified Dok-related protein (DokR) as a substrate for the Src family kinase Lyn. DokR phosphorylation leads to its association with signaling proteins like GAP and Nck, suggesting its role in multiple cellular pathways.

Area of Science:

  • Molecular Biology
  • Cell Signaling
  • Biochemistry

Background:

  • Src family kinases, like Lyn, play crucial roles in cellular signaling pathways.
  • Identifying novel substrates of kinases is essential for understanding signal transduction.
  • Dok-related proteins (DokR) are homologous to known signaling regulators.

Purpose of the Study:

  • To identify novel substrates of the Src family kinase Lyn.
  • To characterize the function and interactions of the identified Dok-related protein (DokR).
  • To elucidate the molecular mechanisms underlying DokR's interactions with downstream signaling partners.

Main Methods:

  • Screening of a murine embryonic cDNA library using a phosphorylation-based strategy.
  • Co-expression of DokR and Lyn in HEK 293T cells and BaF/3 hematopoietic cells.
  • Immunoprecipitation and Western blotting with specific antibodies.
  • Glutathione S-transferase (GST) fusion protein binding assays and site-directed mutagenesis.

Main Results:

  • Dok-related protein (DokR) was identified as a Lyn kinase substrate.
  • DokR is highly expressed in lymphoid tissues.
  • Phosphorylated DokR associates with RasGTPase-activating protein (GAP) and Nck adaptor protein.
  • Specific tyrosine residues (Tyr-276, Tyr-304) mediate DokR binding to GAP, while Tyr-351 mediates binding to Nck.

Conclusions:

  • DokR is a novel substrate of the Lyn kinase.
  • DokR acts as a signaling scaffold, linking Lyn kinase activity to downstream effectors like GAP and Nck.
  • DokR's interactions suggest its involvement in multiple signaling pathways, particularly in lymphoid tissues.

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